Electrochemical glucose test strips, such as those used in the OneTouch® Ultra® whole blood testing kit, which are available from LifeScan, Inc., are designed to measure the concentration of glucose in a blood sample from patients with diabetes. The measurement of glucose is based upon the specific oxidation of glucose by the flavo-enzyme glucose oxidase (GOx(ox)). During this reaction, the enzyme becomes reduced which is denoted as (GOx(red)). The enzyme is re-oxidized by reaction with the oxidized mediator ferricyanide (Fe(CN)63−), which is itself reduced during the course or the reaction. These reactions are summarized below.

When the reaction set forth above is conducted with a test potential applied between two electrodes, an electrical current may be created by the electrochemical re-oxidation of the reduced mediator (ferrocyanide or Fe(CN)64−) at the electrode surface. Thus, since, in an ideal environment, the amount of ferrocyanide created during the chemical reaction described above is directly proportional to the amount of glucose in the sample positioned between the electrodes, the current generated would be proportional to the glucose content of the sample. A mediator, such as ferricyanide is a compound that exchanges electrons between a redox enzyme such as glucose oxidase and an electrode. In a different type of glucose test strip, the enzyme glucose dehydrogenase using a pyrroloquinoline quinone (PQQ) co-factor can be used instead of glucose oxidase. As the concentration of glucose increases, the amount of reduced mediator that is formed also increases, hence, there is a direct relationship between the glucose concentration and the current resulting from the re-oxidation of reduced mediator. In particular, the transfer of electrons across the electrical interface results in a flow of current (2 moles of electrons for every mole of glucose that is oxidized). The current resulting from the introduction of glucose may, therefore, be referred to as an analyte current or more particularly as a glucose current.
Since monitoring of blood glucose levels is an important tool in managing diseases such as diabetes, test meters, using the principles set forth above, have become popular. The glucose current generated during a test is recorded by the test meter and converted into a glucose concentration reading using a preset algorithm that relates current to glucose concentration via a simple mathematical formula. In general, the test meters work in conjunction with a disposable test strip that includes a sample reaction chamber containing electrodes and a reagent. In use, the user deposits a small sample of blood in the sample reaction chamber which is analyzed by the test meter to provide the user with a blood sugar level.
In electrochemical terms, the function of the test meter is twofold. Firstly, it provides a polarizing test potential (e.g., 0.4 V) that polarizes the electrical interface and allows a cell current to flow between two working electrode surfaces. Secondly, the test meter may measure the cell current. The test meter may, therefore be considered to be a simple electrochemical system that operates in a two-electrode mode although, in practice, third and, even fourth electrodes may be used to facilitate the measurement of glucose and/or perform other functions in the test meter.
In most situations, the equations set forth above are considered to be a sufficient approximation of the chemical reaction taking place in the test strip such that a reasonably accurate representation of the glucose concentration is obtained. However, under certain circumstances and for certain purposes, it may be advantageous to improve the accuracy of the measurement, for example, where a portion of the current measured at the electrode results from the presence of other chemicals or compounds in the sample. Where such additional chemicals or compounds are present, they may be referred to as interferents and the resulting additional current may be referred to as an interferent current.
Examples of potential interferents (i.e., compounds found in physiological fluids such as blood that may generate an interferent current in the presence of a test potential) include ascorbate, urate and acetaminophen (Tylenol™ or Paracetamol). A first mechanism for generating an interferent current in a test meter involves the oxidation of one or more interfering compounds by reduction of the mediator (e.g. ferricyanide). In turn, the resulting reduced mediator can then be oxidized at the working electrode. This first mechanism may also be referred to as an indirect interferent current. A second mechanism for generating an interferent current in a test meter involves the oxidation of one or more interferents at the working electrode. The second mechanism may be referred to as a direct interferent current. Thus, the measured cell current includes unwanted contributions from interferents.
A strategy that can be used to decrease the interferent effect is to use a second working electrode in conjunction with a first working electrode and reference electrode. If the second working electrode is bare, then the second working electrode can measure a direct interferent current. The first working electrode should have an enzyme and mediator for measuring a current which includes the summation of a glucose current, a direct interferent current, and an indirect interferent current. The direct interferent current measured at the second working electrode can be subtracted from the current at the first working electrode to reduce the effect of interferents.
Alternatively, or additionally, the second (or third) working electrode can be coated with a mediator (but not an enzyme) to allow the second working electrode to measure a current that includes a summation of the direct and indirect interferent current (but not glucose). In this case, the direct and indirect interferent current measured at the second working electrode can be subtracted from the current at the first working electrode to reduce the effect of interferents.
A disadvantage of using a second (or third) working electrode to compensate for the effects of interferents is that the second working electrode incrementally increases the sample reaction chamber volume and it is preferable that the sample reaction chamber be small so that users do not have to provide a large blood sample. A further disadvantage of using a second working electrode is that it increases manufacturing cost and complexity. Therefore, there is a need to develop methods for measuring glucose independent of interferents using only two electrodes.